1. Field of the Invention
The present invention relates to novel polypeptides capable of binding to the heavy chains (also refered to as .beta.-chains) of the human IL-2 receptor, thereby preventing human IL-2 from binding to the heavy chains of the human IL-2 receptor. The polypeptides according to the present invention can be included in a pharmaceutical compositions and are useful in immunomodulation for e.g. preventing graft rejection or curing autoimmune diseases, or leukemia, and thus they may be used as immunosuppressive agents.
2. Discussion of the Background
Due to the surprising improvement in surgical treatment for organ transplantation, the key to success therein has now been narrowed down to whether the graft rejection after the treatment can be inhibited. Graft rejection is caused when the body recognizes the graft as an extraneous material, and a series of immune responses are elicited to exclude it. Therefore, medicines such as so-called immunosuppresive agents including steroids, azathioprine, methonium compounds, 6-mercaptopurine, etc. have hitherto been administered as graft rejection inhibiting drugs. No significant increase in graft survival rate, however, has been observed because of their narrow safety margins and low effects, etc. Nevertheless with recently developed cyclosporin A the survival rate was drastically improved. Notwithstanding this, the drug was thereafter found to be severely nephrotoxic, and now its use is largely limited.
Accordingly, it has been desired that a less toxic and more effective immunosuppresive agent be developed.
IL-2 is a protein produced by helper T-cells, and has a wide variety of in vivo functions including growth and differentiation induction of killer T-cells and differentiation induction of B cells, thus being a very important factor of defence mechanisms. In organ- and bone marrow transplantation killer T cells, which have been activated by IL-2 or the like, have been shown to play an important role in host versus graft reaction (HVG reaction) or graft versus host reaction (GVH reaction) which is considered to hold the key to successful graft survival.
On the other hand, the immune system of the body is believed to attack the body itself if the system is disturbed, thereby causing autoimmune diseases, the causes for which most likely involve excessive production of IL-2 and other immunological factors or the overresponse thereto.
These facts suggest that selective and effective inhibition of response to IL-2 will lead to prevention of organ transplantation rejection and a cure for autoimmune diseases. In fact, it has been reported that IL-2, which is linked with a cellular toxin capable of selectively injuring IL-2 responsive cells with a receptor for IL-2 (hereunder abbreviated as IL-2 receptor), delayed the onset of and relieved arthritis when it was given to a rat suffering from adjuvant arthritis, an animal model for autoimmune diseases, and further that the immunological rejection by a mouse to the heart transplanted from another mouse of the same species was suppressed with the IL-2 compound administered upon the transplantation (Proc. Natl. Acad. Sci. USA, 86, 1008, 1989).
The IL-2/cellular toxin compound, however, has a short half-life in the blood, and thus its dose must be increased in order to be satisfactorily efficacious, thereby presenting the problem of side effects because of its large dose. In view of these facts, a less toxic medicine has been sought which adequately inhibits the response to IL-2.
The IL-2 receptor on IL-2 responsive cells is known to be composed of two glycoprotein molecules both capable of binding to IL-2 and comprising a .beta. chain having a molecular weight of about 70.000 D and an .alpha. chain whose molecular weight is about 55.000 D.
It has been revealed, that the dissociation constant of the complex between IL-2 and the .alpha. chain is 10.sup.-8 M and for the .beta. chain 10.sup.-9 M respectively, whereas the value decreases to as low as 10.sup.-12 M if both, the .alpha. and the .beta. chains simultaneously bind IL-2 to form a strongly joined three membered associate. Cells with only the .alpha. chain present thereon do not respond to IL-2 bound thereto, but cells with both the .alpha. and .beta. chains present thereon cause various biological activities due to the binding of IL-2. Therefore it is now understood, that the presence of the .beta. chain in the IL-2 receptor is indispensable to the performance of physiological activities of IL-2. That is, it is assumed, that the response to IL-2 will be inhibited by blocking the binding between IL-2 and the .beta. chain.
However, at present there are no substances known, which specifically inhibit the linking between IL-2 and the .beta. chains of IL-2 receptors, with the exception of an anti-IL-2 receptor chain antibody (refer to Japanese Patent Application Disclosure No. HEI 2-18527). Nevertheless this single known antibody is a foreign protein obtained by immunizing xenogenic rabbits and mice. Thus, when the antibody is administered to a human in the form obtained, an immune response occurs against it causing serious side effects such as anaphylactic shock and serum diseases. It is surmised, that the effect of the antibody is weakened, which unfortunately complicates its immediate use in clinical situations.
In order to overcome the drawbacks of these antibodies prepared from xenogenic animals in clinical applications, genetic engineering techniques were developed to convert the constant region (also called C region) of an antibody, which plays no direct part in binding to the antigen to the C region of a human antibody (Methods in Enzymology 178, 459, 1989), heightening hopes for clinically applicable antibodies.
Generally, when an antibody is clinically used, the expected effects of said antibody may be divided into two large categories. The first type of effect is where an antibody combines with an antigen to form an immune complex, and the complement or immune cells are mobilized to eliminate the antigen. This is true for cell surface antigens, etc., in which case the c region of the antibody is indispensable for stimulation of the complement system, and an antibody whose C region is converted to a human type would be effective. The other effect is where the antigen activity is suppressed due to binding of the antibody to the antigen. This includes the case, where the antigen is a bioactive substance or its receptor, or an enzyme, in which case the C region of the antibody is unnecessary. On the contrary the absence of the C region is most desirable, since by administration of the antibody into the body an immune complex thereof with an antigen is formed. In this case, if the antigen is a cell then the cell may be destroyed, and if the antigen is a soluble antigen then it may be trapped in the kidneys causing inflammation. In addition, the molecular weight of the complete antibody containing both the variable regions (also called V region) and the C regions is 150-900 KD compared to a molecular weight of as small as 27 KD for an antibody composed of only the V regions, for which reason the latter is more easily administered to the body.
In order to produce only the V regions of an antibody from antibody protein, it is necessary to obtain a heavy chain (hereunder called H chain) of the antibody and a light chain (hereunder called L chain) of the V region through protease digestion, and induce the binding between them through an appropriate method in order to produce a functional antibody V region molecule. However, it is impossible with present techniques to produce a functional antibody V region molecule with high efficiency and ease.